The overall aim of this research is to identify and characterize gene regulation events that propel rate limiting steps during tumor promotion and tumor progression. The AP-1 transcription factor is a heterodimer of Jun and Fos family proteins that binds to a specific sequence on the transcriptional promoter of certain genes and drives their transcription. Our 1989 observation (Bernstein and Colburn, Science, 1989) that transformation sensitive (P+) but not transformation resistant (P-) mouse JB6 cells responded to tumor promoters by activating AP-1 dependent transcription, suggested that AP-1 activation might be required for progression from preneoplastic to neoplastic(tumor)phenotype. Testing of this hypothesis revealed that the pharmacologic inhibitors, glucocorticoids and retinoids and the "gene therapy" inhibitor dominant negative jun (TAM67) blocked both AP-1 activation and transformation response (Dong et al PNAS, 1994, Li et al Cancer Res. 1996). A second transcription factor NFkappa B was found to be coordinately regulated with AP-1, suggesting the possible importance of both factors in transformation (Li et, Cancer Res 1997). Current research is directed to defining the molecular interactions that regulate AP-1 transrepression by retinoids. The dominant negative jun mutant(TAM67) transgene driven by a keratin 14(K14)promoter, when expressed in a mouse keratinocyte line suppressed both AP-1 and NFkB transcription factor activities as well as induced invasion into matrigel (Dong et al., Molec. Carcinog.,1997)suggesting the possible importance of NFkB in both the cause and prevention of progression. Both AP-1 and NFkB activities and DNA binding show progressive elevation in a human keratinocyte progression model. When the transgene K14-TAM67 or a tetracycline regulated TAM construct was expressed in the more progressed human cell lines that are tumorigenic or anchorage independent, tumor cell phenotype was suppressed (Li et al., Oncogene, 1998 and Li et al., Molec Carcinog. in press). Expression of TAM67 in mouse JB6 P+ cells produced phenotypic reversion to P- phenotype when cells were grown on a nude mouse graft bed (Strickland et al., Carcinog. 1997). Transgenic mice expressing the K14-TAM67 transgene have been generated. In DMBA-TPA initiation-promotion skin carcinogenesis experiments these K14-TAM 67 mice show 90% inhibition of AP-1 activation and 95% protection against promotion of skin carcinogenesis, i.e., prevention of premalignant papilloma formation(Young et al., PNAS, 1999). There is no evidence of toxicity or inhibition of the hyperplasia response to tumor promoter. Recently we have found that the transformation resistance of a JB6 P- variant is due to a shortage of the MAPK kinases Erks 1 and 2 which are also limiting for AP-1 transactivation (Huang, et al, PNAS, 1998). Expression of dominant negative Erk 2 in AP-1 responsive cells has demonstrated the requirement of Erk for AP-1 activation and transformation responses (Watts et al 1998). Current research is directed to identifying Erk dependent molecular events that are required for activating AP-1. Recent observations have identified NFkB non-responsiveness as an explanation for transformation non-responsiveness in the JB6 model(Hsu et al, submitted). P- cells owe their transformation nonresponsiveness to an inability to activate NFkappa B p65 which in turn is attributable to an inability to activate AP-1 dependent transcription. Thus the observation that targeting AP-1 and NFkB elevation prevents tumor promotion and progression has been extended from the mouse JB6 model to mouse and human keratinocyte progression models, and to grafting and transgenic mouse models. New understanding of limiting molecular interactions is emerging. Both the human keratinocytes and the transgenic mice expressing AP-1 inhibitor TAM 67 present valuable opportunities to identify AP-1 target genes whose expression is critical to neoplastic transformation.